Colorectal cancer (CRC) is one of the most common causes of cancer-related deaths in the world. Approximately 1,360,000 new cases were diagnosed globally in 2012, resulting in ˜693,000 annual deaths. These numbers are expected to nearly double over the next 20 years with a rapid rise in obesity and more developing countries adopting a Western diet. Greater focus on early detection of pre-malignant lesions (dysplasia) is needed [Vogelstein et al., Science, 339: 1546-1558 (2013)].
Endoscopy is a frequently performed imaging exam that is widely accepted by patients and referring physicians. However, a significant miss rate of >25% has been found on back-to-back exams for grossly visible adenomatous polyps. Moreover, flat lesions can give rise to carcinoma, and has been found to be as high as 36% of all adenomas. Flat lesions have been found to be more aggressive than polyps, and five times more likely to harbor either in situ or submucosal carcinoma in some patient populations. Studies of outcomes also show that colonoscopy results in a minimal reduction in mortality for cancers that arise in the proximal colon (right side). Furthermore, cancer diagnosed after a “negative” colonoscopy occurs more frequently in the proximal colon. These findings have been attributed to greater genetic instability and a flat morphology. Thus, imaging methods that are sensitive to flat lesions may improve detection and prevention of CRC. Although colonoscopy is widely performed for screening, there is minimal reduction in mortality for carcinomas that arise in the proximal colon. Furthermore, cancer diagnosed after a “negative” colonoscopy occurs more frequently in the proximal colon. These findings have been attributed to greater microsatellite instability and a flat morphology.
Endoscopic imaging with use of exogenous fluorescent-labeled probes, is a promising method for achieving greater specificity in the detection of neoplastic lesions by identifying the expression of unique molecular targets. Imaging provides precise localization, and fluorescence provides improved contrast. Previously, several diagnostic molecules have been used as targeted agents, including antibodies and antibody fragments, for the detection of pre-malignant and malignant lesions in various types of cancer. However, the use of antibodies and antibody fragments is limited by immunogenicity, cost of production and long plasma half-life. Small molecules, RNA aptamers, and activatable probes have also been used. Peptides represent a new class of imaging agent that is compatible with clinical use in the digestive tract, in particular with topical administration.
Phage display is a powerful combinatorial technique for peptide discovery that uses methods of recombinant DNA technology to generate a complex library of peptides, often expressing up to 107-109 unique sequences, that can bind to cell surface antigens. The DNA of candidate phages can be recovered and sequenced, elucidating positive binding peptides that can then be synthetically fabricated. Phage display identified peptide binders to high grade dysplasia in Barrett's esophagus [Li et al., Gastroenterology, 139:1472-80 (2010)] and human colonic dysplasia [Hsiung et al., Nat. Med., 14: 454-458 (2008)] using the commercially available NEB M13 phage system. The T7 system has proven effective in in vivo panning experiments identifying peptides specific to pancreatic islet vasculature [Joyce et al., Cancer Cell, 4: 393-403 (2003)], breast vasculature [Essler and Ruoslahti, Proc. Natl. Acad. Sci. USA, 99: 2252-2257 (2002)], bladder tumor cells [Lee et al., Mol. Cancer Res., 5(1): 11-19 (2007)], and liver tissue [Ludtke et al., Drug Deliv., 14: 357-369 (2007)]. Panning with intact tissue presents additional relevant cell targets while accounting for subtle features in the tissue microenvironment that may affect binding.
Claudin-1 (sometimes abbreviated CLDN1 herein) is an integral membrane protein that forms tight junctions between epithelial cells to regulate paracellular transport [Mrsny et al., Am. J. Pathol., 172: 905-915 (2008)]. This protein is overexpressed in several human cancers, including colorectal [Dhawan et al., J. Clin. Invest., 115:1765-1776 (2005); Miwa et al., Oncol. Res., 12: 469-476 (2001); d Oliveira et al., FEBS Lett., 579: 6179-6185 (2005)], pancreas [Iacobuzio-Donahue et al., Am. J. Pathol., 160: 1239-1249 (2002)], cervical [Lee et al., Gynecol. Oncol., 97: 53-59 (2005)], squamous cell [Morita et al., Br. J. Dermatol., 151: 328-334 (2004)], stomach [Resnick et al., Hum. Pathol., 36: 886-892 (2005)], and thyroid [Fluge et al., Thyroid, 16: 161-175 (2006)]. In colon, claudin-1 is found at levels >40-fold higher in neoplastic by comparison to normal mucosa [Kinugasa et al., Anticancer Res., 27(6A): 3729-3734 (2007)]. These results have been found on gene and protein expression profile analyses [Sheffer et al., Proc. Natl. Acad. Sci. USA, 106: 7131-7136 (2009)], and have been validated on immunohistochemistry [Dhawan et al., J. Clin. Invest., 115: 1765-1776 (2005)]. Overexpression of this protein in neoplasia is believed to increase cell proliferation, motility, and invasiveness, and may contribute to loss of cell polarity, abnormal cellular organization, and decreased differentiation [Gröne et al., Int. J. Colorectal Dis., 22: 651-659 (2007)]. Claudin-1 has also been found to have increased expression in neoplasia associated with inflammatory bowel disease [Mees et al., Int. J. Colorectal Dis., 24: 361-368 (2009); Weber et al., Lab. Invest., 88: 1110-1120 (2008); Kinugasa et al., Anticancer Res., 30: 3181-3186 (2010)].
Various methods for screening for or diagnosing cancer involving claudin-1 are mentioned in WO 2012/135824, WO 2003/069307, US 20080227098, WO 2002/014499 and WO 2012/083338.
New products and methods for early detection of pre-cancer (dysplasia), early cancer and cancer are needed in the art. New products and methods for early detection would have important clinical applications for increasing the survival rate for CRC and other epithelial cell-derived cancers, and for reducing the healthcare costs.